Method For Microbial Control Of Injection Liquid Flow in a Hydrocarbon Reservoir

ABSTRACT

Disclosed herein are methods of establishing a microbial plug in a hydrocarbon-containing geological formation which has been flooded with water. Also disclosed herein are methods of maintaining such plugs and methods of controlling alteration of the position and/or extent of an established plug. Disclosed herein are plugs such as microbial plugs.

CROSS-REFERENCE TO RELATED APPLICATIONS

This application is a continuation of U.S. application Ser. No.14/119,720 filed Nov. 22, 2013, which is a National Phase application ofInternational Application No. PCT/GB2012/051219 filed May 31, 2012,which claims the benefit of priority to Norwegian Patent Application No.20110794 filed May 31, 2011. The content of these earlier filedapplications is hereby incorporated by reference in their entirety.

The invention relates to a method for forming a plug which reduces orprevents flow of liquid in a hydrocarbon-containing geologicalformation. More particularly, position and extent of a plug formed bybiological activity the invention relates to a method for controllingthe in the formation.

It is known in the specialist field in question that liquid can beinjected down into a hydrocarbon reservoir in order to increase thedegree of recovery, especially of oil, from such a reservoir. Injectionliquid is pumped down into the reservoir from one or more injectionwells. The injection liquid is intended to flow through the reservoir asfar as one or more production wells. The injection liquid is intended toform a hydrophilic front which pushes hydrophobic hydrocarbon,especially oil, before it towards the production well. The injectionliquid will also contribute to maintaining the pressure in theformation. The injection liquid will follow a path where the flowresistance is at its least, hereinafter referred to as a flow channel.In the specialist field, this is known as “channelling” or “fingering”.It has been found that use of injection liquid increases the degree ofrecovery, but also that there is still a lot of oil left in thereservoir when the liquid front reaches the production well. The wellwill then produce with too much water mixed in the oil, making furtherproduction unprofitable.

It is possible, to a certain extent, to calculate or model how theinjection liquid flows in the reservoir. For example, it is known forlow-level radioisotopes with relatively short half-lives to be added astracers to the injection water. Radiation from the isotopes will be ableto be identified in the production well. It is thus possible to estimatethe time it takes for the injection liquid to pass through the flowchannel from the injection well to the production well. Alternatively,specific chemicals, for example nitrate, can be used as trace substance.

The ability of the injection liquid to force the oil forwards isreferred to in the field as the sweep efficiency of the injectionliquid. The injection liquid will have this sweep efficiency in the flowchannel, but not in areas that surround the flow channel. It is knownthat the sweep efficiency of the injection liquid can be improved if aplug is formed in the flow channel. The plug can be partially permeable,but the flow resistance increases such that the injection liquid isforced to flow around the plug and thus into those parts of thereservoir that now have the least flow resistance.

The sweep efficiency of the injection liquid is improved in this way.Such plugs can be produced by admixing gel-forming, water-solublepolymers to the injection water. The polymers can be synthetic, forexample polyacrylamide, or biological. Xanthan, for example, is used asa biopolymer and is discussed inter alia in patent documents U.S. Pat.No. 4,716,966, U.S. Pat. No. 4,485,020, U.S. Pat. No. 4,947,932, and GB2,246,586. Patent document U.S. Pat. No. 5,028,344 discusses the use ofcellulose and modified cellulose, while patent document U.S. Pat. No.5,010,954 discusses the use of guar gum and carboxymethylcellulose.

It is also known in the field to add exogenous microorganisms to thereservoir. This is part of what is described as a third-generationtechnique for enhanced oil recovery, in particular for microbialenhanced oil recovery. This is done by mixing microorganisms, andnutrients for the microorganisms, into the injection liquid. Themicroorganisms added can form a biofilm in situ. The microorganisms canalso form biopolymers in situ, for example xanthan. Individually or incombination, cells, biofilm and polymers can form a plug. Patentdocument U.S. Pat. No. 4,799,545 teaches the use of a spore-forming,halo-tolerant, thermo-tolerant and facultatively anaerobic bacterium.The bacterium is added to the injection liquid in spore form and isintroduced down into the reservoir. Strains of Bacillus licheniformisare specifically mentioned as being well-suited for the purpose.Thereafter, sucrose and polyphosphate are added as nutrients to theinjection liquid, and it is then conceivable that B. licheniformis, onaccount of cell growth and the exopolymers formed, will produce a plugin the flow channel. Patent document U.S. Pat. No. 5,174,378 disclosesisolating bacteria that are naturally present in the reservoir. Theseare further isolated by their ability to break down selectedsurfactants. The bacteria can form very small cells when starved,so-called ultramicrobacteria. It is considered that small cells of thiskind can more easily penetrate the pore space of the formation. Asurfactant-containing foam is injected together with the bacteria. Thebacteria return to their active stage, break down the surfactant andproduce exopolymers, such that the flow channel is plugged. Patentdocument U.S. Pat. No. 4,460,043 discloses first adding a suitablebacterium, such as Leuconostoc sp., to a reservoir. The bacterium ispreferably added while it is in a good state of growth, exponentialgrowth, together with suitable nutrients which maintain the growth butwhich do not stimulate the bacterium to produce exopolymers. When thebacterium is established at the desired location in the flow channel,sucrose is added to the injection liquid. For certain types of bacteria,sucrose will act as a stimulant that triggers production of exopolymers.It is also proposed to inject the bacteria from the injection well andsucrose solution from the production well. The plug will then be formedwhere the two streams meet in the reservoir. This will increase thedegree of control over how the plug is formed in the flow channel.Patent document U.S. Pat. No. 4,558,739 discloses injecting a nutrientsolution that includes molasses, grain wort and malt, for stimulatingendogenous bacteria in the flow channel to grow and form, which shouldin itself increase the flow resistance. In addition, the endogenousbacteria will be able to form exopolymers, which will help in the plugformation. It is proposed to stop the injection for a period of 1 to 10days in order to increase the effect of the supply of nutrients. Thepatent document also discloses using strains of Bacillus and Pseudomonasto plug a flow channel. Patent document U.S. Pat. No. 4,475,590discloses stimulating endogenous bacteria in an oil-containing reservoirby adding nitrogen and phosphorus to the injection water. The bacteriause the oil as a carbon source, and the bacteria produce fatty acids andfatty alcohols which reduce the surface tension between the injectionwater and the oil. The bacteria can also produce exopolymers. Asexamples of suitable types of bacteria, mention is made of bacteria ofthe genera Pseudomonas, Achromobacter, Arthrobacter, Flavobacterium,Vibrio, Acinetobacter, Bacillus, Micrococcus and Clostridium. Patentdocument U.S. Pat. No. 4,905,761 shows that it is also known thatinjection of bacteria into a reservoir from an injection well can leadto undesired plugging of the reservoir. The patent document teaches thata microbial formulation comprising types of Bacillus and Clostridiumwill produce a mixture of surfactants, such as glycolipids,lipoproteins, polysaccharide-fatty acid complexes, mono- anddiglycerides and neutral lipids, and solvents, such as short-chainalcohols, ketones and acids. Such a mixture is advantageous forreleasing oil from the reservoir and for transporting oil through thereservoir. The patent document also teaches that types of bacteria ofthe genera Acinetobacter, Arthrobacter, Candida, Corynebacterium,Nocardia, Pseudomonas, Rhodococcus and Toruloosis will be able toproduce surfactants, while types of bacteria of the genera Acetobacter,Arthrobacter, Bifidobacterium, Corynebacterium, Gluconobacter,Lactobacillus, Leuconostoc, Pediococcus, Pseudomonas, Ruminobacter,Ruminococcus, Sporolactobacillus, Streptococcus and Zymomonas will beable to produce solvents.

It is therefore known in the field to increase oil recovery from areservoir by adding nutrients to the injection liquid in order tostimulate endogenous microorganisms to produce biosurfactants andsolvents in situ, and to produce biopolymers so as to form a plug. Aplug can also be formed when microorganisms in situ increase their cellvolume, when supplied with nutrients, and thus block pores in thereservoir. The same effects can also be achieved by adding exogenousmicroorganisms to the injection liquid, either in the form of spores oras active cells.

Nevertheless, known plugging systems are rather limited and are notwidely used in the industry. The present inventor has sought to improveon existing plugging technology. Improvements may include economicbenefits, including the cost of raw materials and enhanced oil recoveryfrom a given reservoir which is in the secondary or tertiary oilproducing phase. It would be highly desirable if a plug was a dynamicsystem, able to respond both to control inputs from the surface and tochanges in the environment in the reservoir. Changes in the reservoirenvironment could include changes in the liquid flow around the plug;for example water which had been forced around a plug in a water channeland into adjacent oil containing regions may clear those areas of oil,enabling the plug to expand into those areas and forcing the water tosweep into new oil containing zones further from the original waterchannel.

After a microbial plug has been established, it may be desirable tochange the position of the plug in the reservoir and to change theextent of the plug in a direction between one or more injection wellsand one or more production wells. It may also be desirable to change theextent of the plug so as to include more than the original flow channel.

It is also desirable to develop suitable alternatives to molasses andsucrose as nutrients for microorganisms in a reservoir. It is alsodesirable to develop a viscous nutrient solution that can also increasethe sweep efficiency of the injection water.

The object of the invention is to overcome or minimize at least one ofthe disadvantages of the prior art, or at least to make available auseful alternative to the prior art.

The object is achieved by the features set forth in the descriptionbelow and in the attached patent claims.

The present inventor has found that by careful selection of themicroorganisms for use in the plug, as well as the nutrients supplied tothem and methods for their introduction, it is possible to provide animproved, dynamic and self-controlling as well as controllable plug.

The invention exploits the fact that microbial growth can be controlledby increasing or decreasing the availability of nutrients, and thatmicroorganisms can be inactivated or killed with the aid of a suitablebiocide. The invention also exploits the fact that the rate of flow ofinjection liquid with nutrients and without nutrients can be increased,decreased or stopped altogether. The invention further exploits the factthat an established plug will lead the injection liquid into geologicalformations that surround the flow channel where a plug is established.

The invention also exploits the fact that the injection liquid is colderthan the petroleum-containing geological formation. This means that,over time, the injection liquid will cool at least some parts of thegeological formation from a temperature that prevents microbial growthto a temperature at which microbial growth is possible. This appliesespecially if both the quantity of injection liquid and also the flowvolume are sufficient. The invention also exploits the fact that a pHregulator can be added to the injection liquid. The regulator can beacidic or basic, and the pH regulator can have buffer capacity. Use isalso made of the fact that a pH regulator in the injection liquid can beused up by reacting chemically with components in the reservoir, suchthat the pH value in the injection liquid will come close to the pHvalue of the reservoir. Especially if the stream of injection liquid isstopped, the pH regulator will diffuse out of the injection liquid andinto the pore water of the reservoir, which has the effect that the pHregulator is diluted and the pH value of the injection liquid, overtime, comes close to the pH value of the reservoir. Use is also made ofthe fact that the injection liquid has, or can have, a lower saltcontent than the pore water in the reservoir. This means that the saltcontent in the reservoir, at least in parts of the reservoir, can bereduced from a concentration that prevents microbial growth to a saltcontent in which microbial growth is possible.

The invention also exploits the fact that micro-organisms can producemetabolites which, in sufficient concentration, have an inhibitingeffect on the growth of the organism. Such metabolites can be externaland/or secondary metabolites. Ethanol is an example of such ametabolite. Use is also made of the fact that gases, for example CO₂,can, at certain concentrations, have a stimulating effect on microbialgrowth. Use is also made of the fact that many microorganisms cannotgrow in the presence of high concentrations of hydrocarbons, especiallyoil, or on substrates covered by hydrocarbons, but that they can growthere when the concentration of hydrocarbons decreases.

The invention also exploits the fact that the thermophilic bacteriumClostridium thermocellum can use cellulose as a carbon source underanaerobic conditions. C. thermocellum has cellulosomes and can breakdown cellulose to cellobiose and cellodextrin, and can break downhemicellulose to xylose, xylobiose and other pentose sugars (Barnard, D.et al., 2010, Extremophiles in biofuel synthesis, Environ. Tech. 31,871-888 (doi: 10.1080/09593331003710236)). Like other Clostridia, C.thermocellum is a spore-forming bacterium.

In the description below, control procedure means any procedure wherethe purpose of the procedure is to act on the microbial activity in apetroleum-containing geological formation. Without being an exhaustivelist and without limiting the invention, a control procedure involves:injecting injection liquid; regulating the quantity and flowrate of theinjection liquid; adding salt, nutrients, pH regulator or biocide to theinjection liquid to an estimated level; regulating the temperature ofthe geological formation with the aid of the injection liquid; inducingmicrobial colonization of surfaces; inducing increased microbialactivity or growth; inducing sporulation in spore-formingmicroorganisms; inducing inactivation or cell death; and inducingproduction of exogenous metabolites, including biofilm, or exogenousmicrobial inhibitors of microorganisms.

In the description below, without limiting the invention, microbialactivity means at least one of the following activities: aerobicrespiration, anaerobic respiration, colonization of surfaces, growth,cell division, sporulation, metabolism, production of endogenous andexogenous primary metabolites, production of endogenous and secondarymetabolites, inactivation and cell death. It will be clear from thecontext which activity is being discussed.

In the description below, microorganisms mean eukaryotic bacteria,archaebacteria and fungi. It will be clear from the context which typeof organism is being discussed. Bacteria and Archaea are preferred.

Preferred properties of the microorganisms are defined in the claims. Inaddition, the microorganisms are preferably facultative anaerobes andare preferably present as a monoculture. The microorganisms (“microbe”is used synonymously with microorganism herein) are typically compatiblewith reservoir conditions in terms of pH and salinity, althoughpreferably the microorganisms thrive at a pH which is different fromnormal reservoir conditions but can be readily applied by the injectionliquid. Preferably the microorganism produces a secreted product whichis inhibitory to growth of the microorganism, e.g. ethanol. While themicroorganism cannot use hydrocarbons as a carbon source, they canpreferably tolerate oil and more preferably oil is not inhibitory togrowth. The microorganisms can preferably grow at a high density andhave a cohesion which means the plug can withstand pressure without theneed for a strong biofilm. The microorganisms preferably exist in sporeform at one or more of, low temperature, starvation and high turbidity,e.g as caused by injection into a reservoir.

The present inventor used a novel set of selection criteria inidentifying suitable microorganisms. In particular it was found thatsuitable candidate microorganisms may be found by selecting for thosespecies which are capable of degrading complex sugars (i.e. using suchsubstrates as a carbon source) and occur naturally in river deltas. Assuch this group represent a preferred class of microorganisms for use inthe methods of the present invention. Either individually or incombination, thermophilicity, oxygen requirements, pH tolerance, abilityto degrade hydrocarbons and tolerance towards hydrocarbons, as well asfactors affecting dormancy and spore formation may also be taken intoconsideration in the selection process.

The microorganisms are preferably cellulolytic and/or hemicellulolytic.In reality, most cellulolytic organisms are also hemicellulolytic. Suchorganisms are able to utilise cellulose or hemicellulose as sole carbonsource. In practice they can also use other carbon sources, inparticular derivatives and degradation products of these complexpolysaccharides, such as the soluble disaccharide cellobiose.

Preferred mircoorganisms according to the invention include Clostridiumthermocellum and Acidothermus cellulolyticus. Different reservoirs areat different temperatures and the microorganisms can be selectedaccordingly; A. cellulolyticus thrives at higher temperatures than C.thermocellum and so is more suitable for hotter reservoirs.

In the description below, injection liquid means injected fresh water orinjected salt water that is supplied to a geological formation throughan injection well. Salt water can include fresh water to which salts areadded, a mixture of sea water and fresh water, natural brackish waterand undiluted sea water. The injection liquid can be degassed,supplemented with biocides or exposed to radiation in order to reducethe number of microorganisms in the injection liquid. Preferably theinjection liquid is less saline than salt water, more preferably thesalinity is 3.5-6%.

In the description below, nutrient solution or growth medium means aninjection solution that has been supplemented with nutrients formicroorganisms. As carbon source or energy source, the nutrients can,for example, contain cellulose, hemicellulose, derivatives of celluloseand hemicellulose, starch, other polysaccharides, oligosaccharides,disaccharides and monosaccharides, a mixture of such saccharides andnutrients containing such saccharides. Preferably the carbon source issoluble. Preferably the growth medium contains a carbon source selectedfrom the group comprising cellulose, hemicellulose, carboxymethylcellulose, cellobiose, xylose, xylobiose and xylan. Cellobiose isparticularly preferred. Sucrose and glucose are generally not suitable.

The nutrient solution can also contain one or more suitable nitrogensources, phosphorus sources, potassium sources and trace elementsources, such as are known in the field. The nutrient solution is agrowth-promoting solution for microorganisms. Suitable components of thegrowth medium, including salts and minerals, are described in theExamples, in particular in Freier medium which represents a suitablegrowth medium. The growth medium is typically mixed with injection waterabove ground and the salinity, pH etc. optimised before injection intothe reservoir.

In a first aspect, the present invention provides a method ofestablishing a microbial plug in a hydrocarbon-containing geologicalformation which has been flooded with water, the method comprising:

-   -   a) introducing into the formation a microbial inoculum, the        microorganisms of which are:        -   (i) spores or otherwise in a dormant state,        -   (ii) capable of sporulation,        -   (iii) cellulolytic or hemicellulolytic,        -   (iv) thermophiles, extreme thermophiles or            hyperthermophiles,        -   (v) unable to utilise hydrocarbons as a carbon source, and        -   (vi) not indigenous to the hydrocarbon-containing geological            formation;    -   b) simultaneously or sequentially introducing into the formation        a growth medium which provides a carbon source which can be        utilised by the microorganisms introduced in step (a) but not by        indigenous microorganisms;    -   c) exposing the inoculum to conditions which enable the        microorganisms to enter an active growth phase within water        channels in the geological formation; and    -   d) introducing an injection liquid comprising further growth        medium as defined in step b) into the formation via an injection        well.    -   The “water channel” may be a void through solid material, akin        to an underground river. Alternatively, a water channel may be a        discrete solid layer which acts as a conduit for water flow        relative to surrounding solid zones of different composition.        Thus, a water channel may be a permeable layer of rock or sandy        material.

Preferably the inoculum is in the form of spores. A dormant state mayconveniently be caused through starvation of the microorganisms. Withoutwishing to be bound by theory, it seems that by introducing the inoculumin a sporous or other dormant state, they are more readily disposed intothe formation, preferably into water channels within the formation. Theinoculum is preferably introduced, together with growth medium, into theformation down an injection well. The flow of liquid into the formationis then reduced or stopped for a period to allow the inoculum to leavethe sporous or dormant state and start to grow. Turbid conditions as arecaused by normal flow rates of injection fluid inhibit growth. Thisperiod of low ‘flow’ or ‘no-flow’ may typically be for 12-72 hours, e.g.around 24 hours, sufficient to allow the microorganisms to enter anactive growth phase. An active growth phase will be defined, forexample, by cell division, increased metabolism and increase in cellsize.

Once active cell growth has begun, the microorganisms can tolerateincreased liquid flow and, indeed, liquid flow is required for the plugto develop fully as further nutrients are required. Continuous liquidflow from an injection well is then preferred for growth and maintenanceof the plug, helping to keep a balance between nutrient input to theplug and removal of any inhibitory products of metabolism such asethanol. The plug has as its primary purpose a reduction in permeabilityin the channel where it grows, however, it is preferably still partiallypermeable, allowing some liquid flow through the plug.

The preferred geological formations are those which contain establishedwater channels before introduction of the inoculum. This canconveniently be determined by a high water content in the output of theproduction well.

In a further aspect, the present invention provides a method ofmaintaining a microbial plug in a hydrocarbon-containing geologicalformation, said plug comprising microorganisms which are:

-   -   (i) capable of sporulation,    -   (ii) cellulolytic or hemicellulolytic,    -   (iii) thermophilic, extreme thermophilic or hyperthermophilic,    -   (iv) unable to utilise hydrocarbons as a carbon source, and    -   (v) not indigenous to the hydrocarbon-containing geological        formation;    -   wherein the formation is flooded with liquid which is injected        into the formation through one or more injection wells and flows        to one or more production wells, the liquid comprising a growth        medium which provides a carbon source which can be utilised by        the microorganisms in the plug but not by indigenous        microorganisms.

It is described later how the plugging system of the present inventionallows for controlled alteration of the plug. More generally, themethods will incorporate phases of maintenance where active steps toalter the position or extent of the plug are not required. However itmay still be beneficial to monitor the plug and the impact it is havingon the reservoir environment. This is conveniently done by monitoringthe output of a production well, for example the oil, water or otherchemical content of the output as well as the rate of output and thetemperature of the output. As a consequence of the composition or otherproperties of the output, the concentration of one or more components inthe injected liquid or one or more physical properties of the injectedliquid may be adjusted. For example, flow rate may be altered, thetemperature of the injected liquid may be changed.

Adjustments to the concentration of components include adding a newcomponent or removing a component entirely. Typically changes will be tothe amount of the component providing the carbon source to themicroorganisms, of other desired nutrients, the salinity of the injectedliquid or its pH.

As demonstrated in the Examples, a preferred feature of the plugs of thepresent invention is that they are removable or substantially removable,for example by cutting off the supply of growth medium. This providesgreater flexibility than, for example, a static plug based on introducedpolymers or secreted products of microorganisms. The biomass itself iscentral to the plugging action according to the present technology andthis provides a dynamic system which can change in situ in response toreservoir conditions and/or to external control stimuli.

pH is a useful tool in the methods of the present invention. Inparticular the pH of the injection liquid containing the growth mediumis preferably selected such that it is inhibitory for indigenousmicroorganisms but the introduced microorganisms can grow at that pH.

The source of carbon which is injected into the reservoir provides asimilar opportunity for preferential growth of the introducedmicroorganisms which form the plug. The source of carbon in the growthmedium is defined herein and is one which cannot be utilised to anysignificant extent, i.e. cannot itself support growth, by the indigenousmicroorganisms who typically use hydrocarbons or glucose or sucrose as acarbon source.

The plug exists, for weeks, months or even longer as a dynamic system inthe formation. The plug creates changes in the hydrocarbon zonessurrounding the water channel in which the plug is established. The plugreceives nutrients in the injected liquid which is preferably providedas a continuous flow, this encourages growth. Growth of the plug islimited by oil in surrounding zones, but as water first drives the oilfrom those areas due to exertion of pressure caused by the presence ofthe plug and is then able to flow into those zones changing then fromoil-wetted zones to water-wetted zones, so the plug can expand intothese areas which are receiving nutrients in the liquid flow.

Temperature is a further self-controlling features to which the dynamicplug responds. Temperatures will increase towards the production wellsand, depending on the microorganism, growth in the flow direction willbe inhibited above a certain temperature. The microorganisms are alsoinhibited by too high a flow rate and this provides a mechanism ofself-adjustment and top-down control. In general, a parabolic shape tothe plug with tails (alternatively considered as a curved funnel whichis partly hollow inside) as shown in the figures, is thought to behighly beneficial and to provide or enhance the functionality of theplugs of the invention; the plugs preferably have such a shape. Theshape may be a function of the availability of nutrients within the plugdue to the semi-permeability thereof and to the production of inhibitorymolecules by the microorganisms.

Preferred microorganisms possess a cellulosome which may contribute tothe permeability and cohesiveness of the plug, as well as to thedigestion of the carbon source.

In a further aspect, the invention relates to a method for controlledalteration of the position and/or extent of an established microbialplug in a hydrocarbon-containing geological formation through which aninjection liquid can flow from one or more injection wells to one ormore production wells, which method comprises selecting a microbialinoculum, an injection liquid and a growth-promoting liquid forformation of a microbial plug, and which method further comprises usingat least one control procedure selected from a group comprising pHregulation, temperature regulation, liquid quantity regulation, gasquantity regulation, inhibitor regulation, nutrient quantity regulation,salinity regulation, viscosity regulation, pressure regulation, flowrateregulation, and addition of further microorganisms to the injectionliquid.

The abovementioned method can be used for controlled movement of theestablished plug in a direction towards at least one production well, inwhich the plug can be formed by at least one type of spore-formingmicroorganism, and which method can comprise:

-   -   using a control procedure to stimulate the microorganism to form        spores;    -   using a control procedure to move the microbial spores in the        geological formation; and    -   using a control procedure to activate the microorganism to break        out of the spores and become active. The control procedure for        stimulating the microorganism to form spores can comprise        reducing the nutrient supply. In an alternative embodiment, the        control procedure for stimulating the microorganism to form        spores can comprise pH regulation of the injection liquid.

The abovementioned method can be used for controlled lengthening of theestablished plug in a direction towards at least one injection well,where the method can comprise:

-   -   the control procedure of supplying fresh nutrients to a plug        front; and    -   the control procedure of reducing the flowrate of the injection        liquid or stopping it completely, such that the microorganism        can grow into a zone containing nutrients between the plug front        and the injection well.

The method can further comprise the control procedure of supplying a gasto the injection liquid in order to stimulate the microbial activity inthe plug front. The gas can be CO₂.

The abovementioned method can be used for controlled expansion of thespread of the established plug at the flanks of the plug, where themethod can comprise:

-   -   the control procedure of supplying fresh nutrients to the        injection water flowing along the flanks of the established        plug; and    -   the control procedure of reducing the flowrate of the injection        liquid or stopping it completely such that the microorganism can        grow into a zone containing nutrients at the flanks of the plug.

The abovementioned method can be used for controlled movement of theestablished plug front in a direction towards at least one of theproduction wells, where the method can comprise:

-   -   using a control procedure to supply biocides to the injection        water to a growth-inhibiting or lethal level.

In an alternative embodiment, the method for controlled movement of theestablished plug front in a direction towards at least one of theproduction wells can comprise:

-   -   using a control procedure to regulate the pH of the injection        water to a growth-inhibiting or lethal pH level such that        microbial activity ceases in the plug front facing towards the        injection well.

The abovementioned methods in the various embodiments in accordance withany of the preceding claims, where the methods can further comprise

-   -   locating an isolated hydrocarbon-containing section in the        geological formation where the permeability is less than the        permeability of the plug and greater than the permeability in an        established flow channel; and    -   positioning the plug to surround the isolated        hydrocarbon-containing section such that the injection liquid        flows through the isolated hydrocarbon-containing section.

In a further embodiment, the method can comprise adding to the injectionliquid, at the desired concentration, biopolymers selected from a groupcomprising cellulose, hemicellulose, derivatives of cellulose, andderivatives of hemicellulose.

The abovementioned regulation of the viscosity can comprise adding tothe injection liquid, at the desired concentration, biopolymers selectedfrom a group comprising cellulose, hemicellulose, derivatives ofcellulose, and derivatives of hemicellulose.

The method described above can further comprise the control procedure ofadding a culture of a strain of Clostridium thermocellum to the freshnutrient liquid.

The method described above can further comprise pretreating the sectionat the plug front or at the flank of the plug by at least one of thefollowing control procedures:

-   -   flooding the plug front or the flank of the plug with injection        liquid containing biocide, in order to reduce the quantity of        active endogenous microorganisms;    -   flooding the plug front or the flank of the plug with injection        liquid containing a pH regulator, in order to improve the growth        conditions for desired microorganisms;    -   flooding the plug front or the flank of the plug with injection        liquid of low salt content, in order to reduce the salinity in        the pore water in the geological formations that surround the        plug front and the flank of the plug, so as to improve the        growth conditions for desired microorganisms; and    -   flooding the plug front or the flank of the plug with injection        liquid in order to cool the geological formations that surround        the plug front and the flank of the plug, so as to improve the        growth conditions for desired microorganisms.

The method described above can further comprise maintaining the shape ofthe plug front, the shape of the flank and the shape of a plug tail byflooding the plug with a nutrient solution, and such that microorganismsin the plug form a microbial inhibitor. The microbial inhibitor cancomprise ethanol.

In a further aspect, the invention relates to a microbial plug formed ina petroleum-containing geological formation, where the plug comprisesmicroorganisms of the type defined above, preferably Clostridiumthermocellum. Preferably the formation is water flooded.

In a further aspect, the invention relates to an injection liquid forflooding a petroleum-containing geological formation, where theinjection liquid contains a polymer selected from a group comprisingcellulose, hemicellulose, derivatives of cellulose, and derivatives ofhemicellulose.

In a further aspect, the invention relates to use of microorganisms ofthe type defined herein, preferably Clostridium thermocellum, forformation of a microbial plug in a petroleum-containing geologicalformation. Preferably the formation is water flooded.

Preferred embodiments are described below by way of example and aredepicted in the attached drawings, in which:

FIG. 1 shows schematically, and seen from above, how an injection liquidflows between an injection well and a production well in a geologicalformation, as is known in the field;

FIG. 2 shows schematically, and seen from the side, ahydrocarbon-containing geological formation where a flow channel isformed between an injection well and a production well, as is known inthe field;

FIGS. 3A-D show schematically and on a microscopic scale: A) howhydrocarbons fill the pore space in a geological formation; B)colonization of the pore space with microorganisms; C) plugging of theformation with microorganisms; and D) shrinkage of the microbial biomassas a result of lack of nutrients, and opening of the pore volume forthrough-flow, as is known in the field;

FIG. 4 shows, at the same scale as FIG. 2, how a microbial plug isformed in the flow channel, as is known in the field;

FIG. 5 shows, at the same scale as FIG. 2, how a type of microbial plugdifferent from that shown in FIG. 4 is formed in the flow channel;

FIGS. 6A-6E show a schematic side view and cross sections of a plug,where the plug's production of inhibiting metabolites leads to a reducedactivity or biomass in the section of the plug towards a productionwell; A) is a schematic representation of the external shape of such aplug 4 seen from the side; B), C) and E) cross sections thatschematically depict the effect of the inhibitor liquid stream 54 on theinternal shape of the plug 4; and D) plug from above;

FIG. 7 shows the extent of a microbial plug towards a production wellafter movement of the plug;

FIG. 8 shows a microbial plug with a greater extent towards the flank ofthe plug;

FIGS. 9A-9D show a highly schematic representation of the phases in themovement of a plug with a direction of flow of injection liquid towardsa production well; A) the established plug 4, and the injection liquidstream 5 that sweeps along the flanks 48 of the plug 4; B) a schematicshowing that spore-forming microorganisms 8, which form the plug 4,sporulate for lack of nutrients; C) how the injection liquid stream 5will move the sporulated microorganisms 8 towards the production well32; and D) how plug 4 is formed; and in

FIGS. 10A-D, A-B are schematic representations of the phases in theelongation of a plug towards an injection well counter to the directionof flow of injection liquid, and Figures C-D show a shortening of theplug's tail.

In the figures, reference number 1 designates a geological formationthat surrounds a hydrocarbon-containing reservoir. The reservoir 1 isdivided schematically into zones: a zone with low permeability 10, azone with greater permeability 12, and a flow channel 2. The flowchannel 2 is formed in a zone where the reservoir 1 has highpermeability and an injection liquid has flowed through the flow channel2. The flow channel 2 is divided into a central zone 20 and a peripheralzone 22. Injection liquid is introduced into the reservoir 1 in a mannerknown per se through an injection well 30. Hydrocarbons and possiblyother liquid is recovered from the reservoir 1 from a production well32. The figures indicate one injection well 30 and one production well32. This is schematic and is to be understood as meaning that there canbe one or more injection wells 30 and one or more production wells 32.An injection liquid stream 5 flows through the flow channel 2. As isshown in FIG. 1, the injection liquid can follow several channels andthe quantity of injection liquid flowing through the flow channel isdependent among other things on the flow resistance. This is shown inFIG. 1 by different widths of the arrows that indicate the injectionliquid stream 5. FIG. 4 shows a microbial plug 4 in the reservoir 1,which plug 4 is formed in a manner known per se.

FIG. 3 shows schematic representations of an assumed mechanism orexplanatory model for drainage of hydrocarbons 7 that are located in apore volume between solid minerals 6, without limiting the invention tothis model. FIG. 3A shows that hydrocarbon 7 fills the pore volumebetween the minerals 6. Many types of microorganisms 8 cannot grow orlive in such hydrocarbon-filled pore spaces. FIG. 3B shows that remainsof the hydrocarbon 7 are still present between the minerals 6 after thereservoir 1 has produced hydrocarbon 7, possibly after injection liquidhas flowed through the reservoir 1 in a known manner. FIG. 3B also showsthat microorganisms 8 are added between the minerals 6. Themicroorganisms 8 can form colonies 82 when they are supplied withnutrient in a nutrient liquid stream 52. The microorganisms 8 will grow,and the colonies 82 will plug the pore volume between the minerals 6, asis shown in FIG. 3C. This will force the injection liquid stream 5 tomove around the plug flanks 48, as is shown in FIGS. 5-9. The reducedflow through the plugged part of the reservoir 1 decreases the nutrientavailability to the microorganisms 8. After the active growth phase, themicroorganisms 8 will enter the terminal phase and die or sporulate ifthe microorganism 8 is a sporulating microorganism 8. The volume of thebiological material decreases, and this part of the reservoir 1 becomesmore permeable, as is shown in FIG. 3D and indicated, for example, inFIG. 5. Residues of hydrocarbon 7 can be released from the minerals 6with the aid of solvents, surfactants and mixtures thereof, and they areconveyed with the injection liquid to the production well 32. Thisincreases the degree of recovery of hydrocarbon 7 from the reservoir 1.Solvents and/or surfactants can be added to the injection liquid.Solvents and/or surfactants can be produced in situ by microorganisms 8in the reservoir 1.

FIG. 4 is a schematic representation of a plug 4 in the flow channel 2.The plug 4 is formed by microbial activity and consists ofmicroorganisms 8 and biopolymers. The plug 4 can be formed by nutrientsin the injection liquid being supplied to the flow channel 2. There willthen be nutrients present along a part of the flow channel 2.Thereafter, exogenous microorganisms 8 in the injection liquid aresupplied to the flow channel 2. Supply of injection liquid is stoppedafter a time, and the microorganisms 8 colonize surfaces in the poresand hollow spaces of the reservoir 1. Since the pores of the reservoir 1will have a certain filter effect, there will be a greater inoculum ofcells at the front 44 of the plug nearer to the injection well 30 inrelation to the production well 32, as is indicated in FIG. 4. Overtime, the microorganisms will spread towards the production well 32since there are nutrients in this area. Production from the productionwell 32 will also contribute to flows in the reservoir 1 towards theproduction well 32. This is indicated in FIG. 4 as a tail 46 in thedirection of the production well 32. The density of microorganisms 8will be greater at the centre 20 of the flow channel 2 than at theperiphery 22. The plug 4 can also be established by injectingmicroorganisms 8 and nutrients at the same time. The plug 4 can also beestablished by first injecting microorganisms 8 and then nutrients. Inthis case, the microorganisms 8 nearest the injection well 30 willconsume nutrients, such that there are fewer nutrients available in theflow channel 2 towards the production well 32. The plug 4 is partiallypermeable, but some of the injection liquid will flow towards theproduction well 32 in the geological formation 1 along flanks 48 of theplug 4. The injection liquid stream 5 will thus carry hydrocarbons withit along the flanks 48 of the plug 4 into peripheral areas 22 of theflow channel 2 and into the geological formation 12 of lowerpermeability.

FIG. 5 shows a microbially formed plug 4 according to the invention,where the plug 4 produces an extracellular inhibitor, which forms aninhibitor liquid stream 54 in the direction towards the production well33. FIG. 6A is a schematic representation of the external shape of sucha plug 4 seen from the side, FIG. 6D shows such a plug seen from above,while FIGS. 6B, 6C and 6E are cross sections that schematically depictthe effect of the inhibitor liquid stream 54 on the internal shape ofthe plug 4. Microorganisms 8 at the flank 48 and tail 46 of the plug 4obtain nutrients, for maintaining their activity, from the nutrientliquid stream 52 that flows along the flank 48 and the tail 46.

FIGS. 9A-D show schematically, in a more simplified form and in moredetail, how the microbially formed plug 4 can be moved towards theproduction well 32, as indicated in FIG. 7. FIG. 9A shows theestablished plug 4, and the injection liquid stream 5 that sweeps alongthe flanks 48 of the plug 4. FIG. 9B shows schematically thatspore-forming microorganisms 8, which form the plug 4, sporulate forlack of nutrients. This is shown as a stippled outline. The permeabilityin the plug 4 increases, and the injection liquid stream 5 flows in theoriginal flow channel 2. The injection liquid stream 5 will move thesporulated microorganisms 8 towards the production well 32, as shown inFIG. 9C. Nutrients are supplied to the injection liquid, and a nutrientliquid stream 52 flows through the flow channel 2 and stimulates themicroorganisms to break out of the spore stage and become active, suchthat a plug 4 is formed as shown in FIG. 9D.

FIG. 8 shows that the flank 48 of the microbially formed plug 4 hasexpanded towards the more hydrocarbon-rich zones 10, 12 in accordancewith the method of the invention. FIG. 10 shows schematicrepresentations of how the front 44 of the microbially formed plug 4 canbe moved towards the injection well 30 in accordance with the method ofthe invention. As is shown in FIG. 10A, a nutrient stream 52 will flowfrom the injection well 30 towards an established plug 4 in accordancewith the invention. Some of the nutrient liquid solution 52 will flowalong the front 44 of the plug 4, while some of the nutrient stream 52will flow through the partially permeable plug 4. The supply ofnutrients maintains the microbial activity of the plug 4. The nutrientliquid solution 52 is stopped, such that there are nutrients in contactwith the front 44 of the plug, and nutrients extend from the front 44 ofthe plug 4 and in the direction towards the injection well 30. The plug4 will form a front 44′, which spreads towards the injection well 30, asis shown in FIG. 10B. FIG. 10C shows that microorganisms 8 in the partof the plug 4 facing towards the production well 32 will decrease inactivity, sporulate or die over time, because of lack of nutrients. FIG.10D shows the shape and position of the active part of the plug 4 aftera period of time. The plug 4 has thus moved to a position closer to theinjection well 4.

EXAMPLE 1

A microbial plug 4 in a permeable geological formation 1 with a flowchannel 2 is formed in a manner known per se, by injecting a nutrientsolution and a bacterial culture into the formation 1 from an injectionwell 30. The nutrient solution and the bacterial culture can be injectedas one mixture in one pulse, and the pulse is followed by injectionliquid without nutrients and bacterial culture. The bacterial cultureconsists of one or more strains of a spore-forming type of bacteria orseveral types of spore-forming bacteria. The bacterial culture canconsist of a strain of Clostridium thermocellum.

Based on the known porosity of the geological formation 1 and the flowcharacteristics in the flow channel 2, it is possible to calculate whenthe mixture of nutrient solution and bacteria 8 will reach the part ofthe flow channel 2 where it is desirable to establish a plug 4. Theinjecting of injection liquid is halted such that the bacteria 8 areable to establish themselves. The bacteria 8 will use the nutrients forcell division and for general metabolism, which also includes formationof exogenous products, for example a biofilm, and for formation ofsecondary metabolites, which can include alcohols, aldehydes, ketonesand gases. The bacteria 8 will grow in that part of the flow channel 2where nutrients are available. Facultatively anaerobic bacteria 8 willform an anaerobic environment and are not dependent on the presence ofoxygen.

The increased number of bacteria 8, possibly together with biofilmproduction, will reduce the permeability of the flow channel 2, as isknown in the field and is shown in FIG. 3. When the plug 4 is consideredto be established, the injecting of injection liquid is recommenced. Atthe front 44 of the plug 4, that is to say the part of the plug 4 facingtowards the injection well 30, the injection liquid will flow mainlyaround the front 44 of the plug 4 and along the flanks 48 of the plug 4where the permeability is greater than inside the plug 4. This has theeffect that the injection liquid entrains hydrocarbons 7 into parts ofthe formation 1 that lie outside the original flow channel 2, as isshown in FIGS. 5-9.

Without supply of fresh nutrients, the bacteria 8 will stop growing andwill gradually sporulate as shown in FIGS. 3D, 9B and 10C. The sporesare much smaller than the active cells and do not plug the pores in thegeological formation 1 in the same way. The pressure applied by theinjection liquid against the front 44 of the plug 4 will press or washthe spores along the flow channel 2 such that they penetrate inside theformation 1, which leads to the front 44 of the plug 4 being movedtowards the production well 32, as shown in FIG. 9C. A plug 4 that hasbeen established by injection of a mixture of nutrients andmicroorganisms will have approximately the same nutrient status over thewhole extent of the plug 4. When the supply of nutrients is stopped,sporulation will take place more or less simultaneously over the wholeplug 4. The front 44 and tail 46 of the plug 4 are in fluidcommunication, such that the spores in the tail 46 of the plug 4 willalso penetrate inside the formation 1 when the spores at the front 44 ofthe plug 4 begin to penetrate inside. Nutrients are added to theinjection liquid, which now once again flows mainly through the flowchannel 2, and the nutrients will cause the microorganisms 8 to breakout of the spores and form active cells. This has the effect that thewhole plug 4 is moved towards the production well 32.

In another embodiment, as an alternative or addition to inducingsporulation by starving, the injection liquid can be provided with acidsor bases, which respectively acidify and raise the pH of the injectionliquid to a level that induces sporulation. Sporulation will then startat the front 44 of the plug 4 and, as the sporulation begins, theinjection liquid will pass further into the original plug 4 and lead tofurther sporulation. At the same time, the part of the plug 4 containingactive microorganisms 8 will counter the flow of the injection liquid inthe flow channel 2. The injection liquid is thus forced out along theflanks 48 to the remaining part of the plug 4 and also carries with it,to the flanks 48, spores from the front 44 of the plug. Theconcentration of the pH regulator of the injection liquid will decrease,because of dilution and chemical reactions, to a level that does notalter the activity of the cells. Thus, the cells in the tail 46 of theplug 4 will not sporulate. By adding nutrients to the injection liquid,the spores in the front 44 and flank 48 of the plug 4 will break out ofthe spores and form active cells. This has the effect that the front 44of the plug 4, but not the tail 46, is moved closer to the productionwell 32, and that the front 44 of the plug 4 is expanded in theperipheral direction.

In an alternative method for moving a plug 4 in the direction from theinjection well and towards the production well 32, the nutrient solutionis injected first, and the microorganisms 8 are injected immediatelythereafter, such that there is contact between the injection liquid withnutrients and the injection liquid with microorganisms. When theinjection is stopped, the microorganisms will spread inside the flowchannel 2 where there are nutrients. The microorganisms 8 will starveand will sporulate in those parts of the injection channel 2 where thenutrients have been used up. This has the effect that the whole plug 4moves towards the production well 32 over time.

In another alternative method, a suitable biocide, for example ammonia,is added in a suitable quantity to the injection liquid. This will firstinactivate or kill the microorganisms 8 in the front 44 of the plug 4.The effect of the ammonia will abate over time as it reacts with theminerals 6 in the reservoir 1 and is diluted. The result is that thefront 44 is moved in the direction towards the production well 32. Themicroorganisms 8 in the tail 46 of the plug will continue growing andwill spread along the flow channel 2 towards the production well 32.

EXAMPLE 2

A microbial plug 4 is established in the manner described in Example 1.After the plug 4 is established, a fresh solution of nutrients isinjected as far as the plug front 44. The pumping-in of the injectionliquid is stopped when the nutrient solution is in contact with the plugfront 44. The bacteria 8 in the plug 4 will grow into the zone withfresh nutrient solution and move the plug front 44 towards the injectionwell 32, as is shown in FIG. 10B, such that a new plug front 44′ isformed. The original plug 4 is partially permeable, such that aproportion of the fresh nutrient liquid stream 52 will penetrate intothe original plug front 44 and onwards to the tail 46 of the plug.However, the nutrients will be consumed in the plug 4, such that theoriginal front 44 of the plug 4 is maintained, while little or no freshnutrients will reach the tail 46 of the plug, and microorganisms 8 inthe tail 46 will become inactive, sporulate or die, as shown in FIGS.10C and 10D. Alternatively, a mixture of microorganisms 8 and freshnutrients can be injected until it is in contact with the plug front 44.A new plug front 44′ will then be established close to the injectionwell 30. When the new plug front 44′ is established, injection liquid isinjected and will entrain hydrocarbons 7 around the plug 4. As thenutrients are used up, microorganisms 8 in the tail 46 of the plug 4facing towards the production well 32 will become inactive, sporulate ordie and will reduce the extent of the plug 4 in this direction. This hasthe effect that the plug 4 is moved towards the injection well 30 overtime.

In an alternative embodiment, CO₂ can be added to the injection liquid,which is pulsed forwards to the plug 4. CO₂ will stimulate growth ofcertain types of bacteria, for example C. thermocellum.

In an alternative embodiment, the direction of flow of the injectionliquid is reversed by means of injection liquid being injected from theproduction well 32 towards the injection well 30. The method is then thesame as described in Example 1.

EXAMPLE 3

A microbial plug 4 is established in the manner described in Example 1.The injection liquid will carry hydrocarbons 7 from the geologicalformations 1 along flanks 48 of the plug 4. When these formations 1,which surround flanks 48 of the plug 4, are drained of hydrocarbons 7and become more permeable and are more easily colonized bymicroorganisms 8, nutrients are added to the injection liquid. When theinjection liquid with nutrients surrounds at least parts of the flanks48 of the plug 4 (see FIG. 10A for example), the injection is stopped.Microorganisms 8 from the plug 4 will grow into the zones with freshnutrient solution and extend the range of the plug 4 along flanks 48 ofthe plug 4. The injection is then started up again, and the injectionliquid will be forced into zones surrounding the extended plug andcontaining hydrocarbons, such that these are swept out as shown in FIG.8.

In an alternative embodiment, microorganisms 8 and nutrients are addedto the injection liquid flowing past the flanks 48 of the plug 4. Whenthe injection liquid with nutrients surrounds at least parts of theflanks 48 of the plug 4, the injection is stopped. This has theadvantage that the expansion of the flanks 48 of the plug 4 takes placemore rapidly.

The fresh nutrients in the injection liquid surrounding the plug 4 alongthe flank 48 and the tail 46 will partially diffuse into the establishedplug 4 and will contribute to the latter being maintained in theseparts.

When the plug 4 is well established, the injection liquid stream 5 willflow past the plug 4 along the plug flank 48 and will sweep hydrocarbons7 towards the production well 32, as is shown in FIG. 8. This means thatthe flow channel 2 is expanded. By adding nutrients to the injectionliquid and, after a suitable period of time, stopping the supply ofinjection liquid, microorganisms 8 in the plug 4 will grow into thezones 10, 12 flanking the original flow channel 2.

EXAMPLE 4

A microbial plug 4 is established in the manner described in Example 1.Injection liquid flows along the plug flank 48 and the tail 46 and, overtime, will sweep out hydrocarbons 7 in the geological formations thatsurround the plug 4. The injection liquid will also cool thesesurrounding geological formations in the reservoir 1. Draining the poresof hydrocarbon 7, in combination with cooling, makes it possible for themicroorganisms 8 to colonize areas around the tail 46 and lengthen thetail 46 towards the production well 32 when nutrients are supplied tothe injection liquid and these nutrients flow past the tail 46 towardsthe production well 32. The plug 4 is maintained by means of themicroorganisms 8 undergoing alternating phases of growth and shrinkage,as has been described above in connection with FIG. 3. In the shrinkagephase (see FIG. 3D), nutrients can be supplied to the plug 4, and themicroorganisms 8 are stimulated to new growth, as shown in FIG. 3C. Thishas the effect that the plug front 44 is kept stationary, while theextent of the plug 4 towards the production well 32 is lengthened.

EXAMPLE 5

A microbial plug is established in the manner described in Example 1,and the front of the plug is moved towards the injection well, asdescribed in Example 3. In addition, the extent of the plug towards theproduction well is maintained by pulsing nutrients into the plug 4, asdescribed above, in order to maintain the microbial activity in the partof the plug 4 facing towards the production well 32. The shape of theplug 4 is thus maintained as shown schematically in FIG. 10B.

In FIGS. 5-10, the plug 4 is shown schematically with a hollow portionbrought about by in situ production of inhibitors. C. thermocellum is anexample of a microorganism 8 that produces ethanol. The ethanolconcentration will increase through the plug and will, above a givenlevel, have an inhibiting effect for microorganisms 8 downstream. Thesewill be inactivated, sporulate or die. This has the advantage that theoutside of the plug 4 (front 44, flank 48 and tail 46) will keep itsposition at the same time as the active volume of the plug 4 is reducedin proportion to a “compact” plug 4. The requirement of the plug 4 fornutrients is therefore reduced. As solvent, ethanol will also contributeto washing out hydrocarbons 7 in the flow channel 2 downstream of theplug 4. Other metabolic products such as acetate and lactate will alsocontribute to increased washing-out of hydrocarbons 7 and will increasethe degree of production from the reservoir 1.

EXAMPLE 6—A LABORATORY SCALE EXPERIMENT WAS PERFORMED TO SHOW PLUGGINGOF A SANDPACK IN ACCORDANCE WITH THE TECHNIQUES OF THE PRESENT INVENTION

Materials and Methods

Bacteria

Clostridium thermocellum (CT) JW20; ATCC 31549

Growth Medium

CT were cultured according to the methodology described by Freier et al.in Applied and Environmental Microbiology [1988] vol 54, No. 1, p204-211.

Specifically, the culture medium contained (per liter of deionizedwater)

1.5 g KH₂PO₄

4.2 g Na₂HPO₄.12H₂O

0.5 g NH₄Cl

0.5 g (NH₄)₂SO₄

0.09 g MgCl₂.6H₂O

0.03 g CaCl₂

0.5 g NaHCO₃

2 g of yeast extract

0.5 ml of vitamin solution. The vitamin solution contained (per liter ofdistilled water)

40 mg of biotin, 100 mg of p-aminobenzoic acid, 40 mg of folic acid, 100mg of pantothenic acid calcium salt, 100 mg of nicotinic acid, 2 mg ofvitamin B12, 100 mg of thiamine hydrochloride, 200 mg of pyridoxinehydrochloride, 100 mg of thioctic acid, and 10 mg of riboflavin.

5 ml of mineral solution. The mineral solution contained (per liter ofdistilled water)

1.5 g of nitriloacetic acid, 3 g of MgSO₄-7H₂0, 0.5 g of MnSO₄. H20, 1 gof NaCl, 0.1 g of FeSO₄ 7H20, 0.1 g of Co(NO₃)₂.6H₂0, 0.1 g of CaCl₂(anhydrous), 0.1 g of ZnSO₄.7H₂0, 50 mg of NiCl₂, 10 mg of CuSO₄*5H₂0,10 mg of AlK₂(SO₄)₃ (anhydrous), 10 mg of boric acid, 10 mg ofNa₂MoO₄*2H₂0, 10 mg of Na₂WO₄-2H₂0, and 1 mg of Na₂SeO₃ (anhydrous).

1% cellobiose

Test Set Up

The apparatus was set up to be able to flow growth medium through asandpack of sandstone size Ø 1-2 mm. A pump was required to deliverpressure up to 20 Bar at maximum head and minimum or zero flow.

The sandpack consisted of a steel 3 m pipe of 1 inch diameter packedwith sand—main chemical component SiO₂. Upstream of the pump a tankcontaining dissolved growth media was installed able to maintain 75° C.and to boil. Between the pump and the inlet to the pipe is a return lineback to the tank which can reduce flow rate through the pipe. A furthervalve after the end of the pipe can also be used to adjust flow rate andsimulate reservoir conditions with respect to pressure.

Water source: spring water boiled for 1 hour. The detailed chemicalcontent of the water is unknown but the water is characterized as hard,ie containing Calcium. The parts from the tank to the output of the pipewere isolated.

Preparation of the Test System

The system was cleaned by boiled water circulating in the system for 3hours and de-oxygenated by N₂-bubbling. The temperature was lowered to75° C. in the tank and re-circulated for 1 week. The system was nowconsidered to maintain a steady state with respect to any microbialpollution. Growth medium was circulated for an additional 3 weeks tolook for any physical behaviour or change in the packed sand section dueto the growth medium.

Observations Prior to Introducing Bacteria:

No pressure built up or change was detected as a result of floodinggrowth medium through the system over time.

Addition of Bacteria:

A small culture of CT bacteria was inoculated and set to grow in a flaskas described in Freier et al. supra, in the growth medium describedabove. The bacteria were then inserted into the sandpack by opening thepipe near the pump end and infecting a small area. The system foropening was a flanged system.

Analysis

Evaluation was based on visual observations and distillation of waterwhich had passed through the pipe to determine the concentration ofethanol per water unit. Visual observation of gases by testing theflammability of H₂ and detection of CO₂ concentration were conducted.Gas tests were performed to confirm that the correct pathway wasfollowed. The concentration of ethanol was used as indication ofefficiency of bacterial (CT) catabolic processes.

Test Case 1.

The manometer read 10 Bars at the beginning of the test.

Active bacteria were introduced into the sandpack within the growthmedia. The media was left to grow for 24 hours and a water flow rate ofmax 20×10⁻⁶ m/s was introduced and the sandpack flooded. This low flowwas maintained and adjusted over 2 weeks.

Observation:

After 24 hours the flow rate was reduced. By reducing flow rate usingthe return line (reject valve), a flow rate of 10×10⁻⁶ m/s wasestablished. The readings on the manometer with closed reject valve (nowater using the return line) showed 20 Bars.

The water penetrating trough the sandpack was reduced dramatically, byapprox 85-95% and the concentration of ethanol increased 2-fold.

Clear indications are that the flooded sandpacks were blocked. Ethanolproduction and gas production indicated active, an Cellulosome-pathway.

Test Case 2:

The same test as described in Test case 1 was continued for one month.

The process was in a steady state: No change was observed in pressure,it stayed at approx 20 Bar and there was no change in flow rate orethanol-production. These results indicate that the plug was maintained.

Test Case 3:

The cellobiose was removed from the growth medium and the system floodedover 7 days.

The pressure decreased and flow rate increased through the sandpack. Thepressure dropped to approx 16 Bar. Changes mainly took place in thefirst 24 hours, from when the changes levelled out to a new steady statesituation. Thus, permeability of the plug is increased but some plugstructure remains.

Test Case 4:

The complete growth medium was removed and only water provided to floodthe system.

The pressure dropped towards 11-12 Bar over 5 days. This suggests theplug is removed/lost or at least that permeability levels return almostto the pre-plug levels.

Test Case 5:

Test case 1 was repeated but with a pipe of 16 inches in diameter.

The results from test 1 were repeated. In this test, a parabolic shapewas traced in the sand when opening the sandpack.

Test Case 6:

Test one was repeated and a plug was established. The growth medium wasremoved and increased flow was introduced until sporous conditions wereassumed to have been established. Maximum flow was introduced into thepipe to flush spores in the flow direction. The spores later started togrow and the centre of the plug was observed to settle in the systemfurther into the.

To finish the test, highly alkaline liquid was flushed through thesystem for 2 days and growth medium was removed. The plug was removed bythis treatment.

EXAMPLE 7

Tests were performed in two different setups, one test containing sandinfected by oily residues and one test only with sand to investigatebacterial growth in a solid environment.

Test One:

Setup with sand only. The sand was of the same type as described inExample 6. The biological input was based on the Freier tests referencedin Example 6. The water used was taken from the same source and treatedthe same way, boiled and de-oxygenated. The CT bacteria were grown inbottles and injected into a 1 litre vessel.

The vessel was completely filled with growth medium and bacteria evenlydistributed in the medium. The flow was then stopped. Temperature wasmaintained at approx 60° C.

The microbes grew and catabolised. We could clearly see that themicrobes thrived and grew in the sand and by measuring the ethanolcontent, the tests confirmed metabolic activity.

The Chemostat-test became a static test for 7 days. After 7 days we sawa change in density and noted the colour to be a little darker. Wepresumed that nutrients were used up and the bugs were in a state ofstarvation. By flushing growth medium through the sandpack system wewere able to change back to a yellowish colour and ethanol was detected.

Test Two:

This test was to establish an environment similar to an empty oilreservoir. This means a sandpack containing 9-16% of oily residues wherethe residue is mainly located on the lee-side of the sand particlesrelative to the flooding direction.

A column with sand according to test one was set up. The column wassaturated with crude oil from the Statfjord field and flooded withseawater. As a result of the flooding approx 88% of the oil wasdisplaced and the system became water saturated. The system was furtherflooded 5 times with growth medium and pre-grown CT culture wasintroduced evenly in the test setup.

The test was set to grow for a week. Corresponding ethanolconcentrations to volume of growth medium were measured.

A habitat contaminated by crude oil did not limit the growth andcatabolic activity of the culture. It seems the oily parts only displacegrowth. Therefore growth and metabolic activity can be maintained sideby side with an oily environment in a microscale system.

EXAMPLE 8

The Freier approach to CT culturing was modified with bottles containing3 different fractions of oil. One set of bottles contained 90% of oil,one set of bottles contained 50% of oil and the last set of bottlescontained 10% of oil. The remaining liquid contained the Freier mediumwith cellobiose at 1%. Bottles containing 100% Freier medium withcellobiose (1%) were provided as control. The bottles were shaken every3 hours during the day. The bottles were opened after 1 week.

Ethanol was produced in concentrations corresponding to the volume andconcentration of growth medium.

The oil did not have a inhibiting effect on the culture. The culturedoes not grow and metabolize within the oily fraction. We concluded thatthe growth medium was effectively removed in the high concentration ofoil due to the fact that oil and water are not soluble. We alsoconcluded that the culture could not utilize Hydrocarbons ascarbon-source.

1. A method of establishing a microbial plug in a hydrocarbon-containinggeological formation which has been flooded with water, the methodcomprising: a) introducing into the formation a microbial inoculum, themicroorganisms of which are: (i) in a dormant state, (ii) cellulolyticor hemicellulolytic, (iii) thermophiles, extreme thermophiles orhyperthermophiles, (iv) unable to utilise hydrocarbons as a carbonsource, and (v) not indigenous to the hydrocarbon-containing geologicalformation; b) simultaneously or sequentially introducing into theformation a growth medium which provides a carbon source which can beutilised by the microorganisms introduced in step (a) but not byindigenous microorganisms; c) exposing the inoculum to conditions whichenable the microorganisms to enter an active growth phase within waterchannels in the geological formation; and d) introducing an injectionliquid comprising further growth medium as defined in step b) into theformation via an injection well.
 2. The method of claim 1, wherein theinoculum is in the form of spores.
 3. A method of maintaining amicrobial plug in a hydrocarbon-containing geological formation, saidplug comprising microorganisms which are: (i) cellulolytic orhemicellulolytic, (ii) thermophilic, extreme thermophilic orhyperthermophilic, (iii) unable to utilise hydrocarbons as a carbonsource, and (iv) not indigenous to the hydrocarbon-containing geologicalformation; the method comprising: a) flooding the formation with liquidby injecting the liquid into the formation through one or more injectionwells, the liquid comprising a growth medium which provides a carbonsource which can be utilised by the microorganisms in the plug but notby indigenous microorganisms, and b) continuing the flow of the liquidthrough the hydrocarbon containing geological formation so that itreaches one or more production wells, thereby maintaining the microbialplug in the hydrocarbon-containing geological formation.
 4. The methodof claim 3, further comprising the step of monitoring the output of aproduction well and as a consequence of the properties of the output,adjusting either the concentration of one or more components in theinjected liquid or one or more physical properties of the injectedliquid.
 5. A method for altering in a controlled manner the positionand/or extent of an established microbial plug in ahydrocarbon-containing geological formation through which an injectionliquid can flow from one or more injection wells to one or moreproduction wells, wherein the microorganisms of the microbial plug are:(i) cellulolytic or hemicellulolytic, (ii) thermophilic, extremethermophilic or hyperthermophilic, (iii) unable to utilise hydrocarbonsas a carbon source, and (iv) not indigenous to thehydrocarbon-containing geological formation; the method comprising: a)selecting an injection liquid and a growth medium which provides acarbon source which can be utilised by the microorganisms of themicrobial plug but not the indigenous microorganisms; and b) applying atleast one control procedure to said plug using said injection liquid,said control procedure selected from a group comprising pH regulation,temperature regulation, liquid quantity regulation, gas quantityregulation, inhibitor regulation, nutrient quantity regulation, salinityregulation, viscosity regulationpressure regulation, flowrateregulation, and addition of further microorganisms to the injectionliquid; thereby altering in a controlled manner the position and/orextent of the established microbial plug.
 6. The method of claim 5 forcontrolling the movement of the established plug in a direction towardsat least one production well, in which the plug is formed by at leastone type of microorganism as defined in claim 3, wherein themicroorganism is spore forming, and which method comprises: using acontrol procedure to stimulate the microorganism to form spores; using acontrol procedure to move the spores in the geological formation; andusing a control procedure to activate the microorganism to break out ofthe spores and become active.
 7. The method of claim 6, in which thecontrol procedure for stimulating the microorganism to form sporescomprises reducing the nutrient supply.
 8. The method of claim 5 forcontrolling the length of the established plug in a direction towards atleast one injection well, which method comprises: the control procedureof supplying fresh nutrient to a plug front such that the microorganismcan grow into a zone containing nutrient between the plug front and theinjection well.
 9. The method of claim 5 for controlling the expansionof the spread of the established plug at flanks of the plug, whichmethod comprises: the control procedure of adding fresh nutrient to theinjection water flowing along the flanks of the established plug suchthat the microorganism can grow into a zone containing nutrient at theflanks of the plug.
 10. The method of claim 5 for controlling themovement of the established plug front in a direction towards at leastone of the production wells, which method comprises: using a controlprocedure to supply biocides or other toxic agent to the injection waterto a growth-inhibiting or lethal level.
 11. The method according toclaim 8 or 9, which method further comprises pretreating the section atthe plug front or at the flank of the plug by at least one of thefollowing control procedures: flooding the plug front or the flank ofthe plug with injection liquid containing biocide, in order to reducethe quantity of active endogenous microorganisms; flooding the plugfront or the flank of the plug with injection liquid containing a pHregulator, in order to improve the growth conditions for desiredmicroorganisms; flooding the plug front or the flank of the plug withinjection liquid of low salt content, in order to reduce the salinity inthe pore water in the geological formations that surround the plug frontand the flank of the plug, so as to improve the growth conditions fordesired microorganisms; and flooding the plug front or the flank of theplug with injection liquid in order to cool the geological formationsthat surround the plug front and the flank of the plug, so as to improvethe growth conditions for desired microorganisms.
 12. The method ofclaim 1 or claim 3, wherein the microorganisms are cellulolytic.
 13. Themethod of claim 12, wherein the microorganisms are bacteria or Archaea.14. The method of claim 13, wherein the microorganisms are Clostridiumthermocellum.
 15. The method of claim 1 or claim 3, wherein the growthmedium contains a carbon source selected from the group comprisingcellulose, hemicellulose, carboxymethyl cellulose, cellobiose, xylose,xylobiose and xylan.
 16. The method of claim 15, wherein the growthmedium contains cellobiose.
 17. A microbial plug formed in a waterflooded petroleum-containing geological formation, characterized in thatthe plug comprises microorganisms as defined in claim
 3. 18. Use ofmicroorganisms of the type Clostridium thermocellum for formation of amicrobial plug in a water-flooded petroleum-containing geologicalformation.
 19. A method of recovering hydrocarbon from a hydrocarboncontaining geological formation wherein said method comprises performinga method as claimed in claim 1 or claim 3 and then recoveringhydrocarbon from said formation.
 20. The method of claim 19, wherein thehydrocarbon is oil.